Biosynthetic studies on terpenoids produced by Actinomycetes by focusing on the nonmevalonate pathway

以非甲羟戊酸途径为重点的放线菌产生的萜类化合物的生物合成研究

• 批准号: 10460047
• 负责人: SETO Haruo
• 金额: $ 4.8万
• 依托单位: The University of Tokyo
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1998
• 资助国家: 日本
• 起止时间: 1998 至 1999
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-10460047/
• 关键词: 3-hydroxy-3-methylglutaryl coenzyme A reductase; 2-C-methyl-D-erythritol 4-phosphate; 4-(cytidine 5'-diphospho)-2-C-methyl-D-erythritol; 2-phospho-4-(cytidine 5' -diphospho)-2-C-methyl-D-erythritol; 2-C-methyl-D-erythritol 2,4-cyclodiphosphate; 1- de

We have obtained the following results.1. Labeling experiments proved that novobiocin produced by Streptomyces niveus was biosynthesized by the nonmevalonate pathway.2. The terpene moiety of BE-40644 produced by Actinoplanes sp. was biosynthesized by the mevalonate pathway, but the cell membrane component, menaquinone, was produced by the nonmevalonate pathway.3. HMG CoA reductase was purified from Streptomyces sp. CL190aeriouvifer and its relevant gene was cloned.4. The key enzyme of the nonmevalonate pathway, 1-deoxyxylulose 5-phosphate reductoisomerase, was purified from E. coli.5. A biosynthetic intermediate of the nonmevalonate pathway, 2-C-metllyl-D-erythrito1 4-phosphate, was converted to 4-(cytidine 5'-diphospho)-2-C-methyl-D-erythrito1 by a new enzyme, 2-C-methyl-D-erythritol 4-phosphate cytidylyltransferase.6. The above mentioned 4-(cytidine 5'-diphospho)-2-C-methyl-D-erythritol was converted to its 2-phosphate derivative by a new enzyme, 4-(cytidine 5'-diphospho)-2-C-methyl-D-erythritol kinase.7. The above mentioned phosphate derivative was converted to 2-C-methyl-D-erythritol 2, 4-cyclodiphosphate.

我们得到了以下结果。1.标记实验证明，雪白链霉菌产生的新生霉素是通过非甲羟戊酸途径生物合成的.游动放线菌产生的BE-40644的萜烯部分是通过甲羟戊酸途径生物合成的，但细胞膜成分甲萘醌是通过非甲羟戊酸途径产生的。3.从产气链霉菌CL 190中分离纯化HMG CoA还原酶，并克隆其相关基因.从大肠杆菌中分离纯化了非甲羟戊酸途径的关键酶1-脱氧木酮糖5-磷酸还原异构酶。coli.5。非甲羟戊酸途径的生物合成中间体2-C-甲基-D-D-胆甾醇4-磷酸被一种新的酶2-C-甲基-D-D-胆甾醇4-磷酸胞苷酰转移酶转化为4-（胞苷5 '-二磷酸）-2-C-甲基-D-D-胆甾醇。上述4-（胞苷5 '-二磷酸）-2-C-甲基-D-腺苷酸被一种新的酶4-（胞苷5'-二磷酸）-2-C-甲基-D-腺苷酸激酶转化为它的2-磷酸衍生物。将上述磷酸酯衍生物转化为2-C-甲基-D-β-2，4-环二磷酸酯。

项目成果

S.Takahashi, T.Kuzuyama, H.Watanabe and H.Seto: "A novel 1-deoxy-D-xylulose 5-phosphate reductoisomerase catalyzing the formation of 2-C-methyl-D-erythritol 4-phosphate in an alternative non-mevalonate pathway for terpenoid biosynthesis." Proc.Natl.Acad.S

S.Takahashi、T.Kuzuyama、H.Watanabe 和 H.Seto：“一种新型 1-脱氧-D-木酮糖 5-磷酸还原异构酶，可催化 2-C-甲基-D-赤藓糖醇 4-磷酸的形成，以替代非

T. Kuzuyama, S. Takahashi and H. Seto: "Construction and characterization of Escherichia coli disruptants defective in the yaeM gene"Biosci. Biotech. Biochem. 63. 776-778 (1999)

T. Kuzuyama、S. Takahashi 和 H. Seto：“yaeM 基因缺陷型大肠杆菌破坏株的构建和表征”Biosci。

T. Kuzuyama, M. Takagi, S. Takahashi and H. Seto: "Cloning and characterization of 1-deoxy-D-xylulose-5-phosphate synthase from Streptomyces sp. strain CL190 that uses both the mevalonate and nonmevalonate pathways for isopentenyl diphosphate biosynthesis

T. Kuzuyama、M. Takagi、S. Takahashi 和 H. Seto：“来自链霉菌菌株 CL190 的 1-脱氧-D-木酮糖-5-磷酸合酶的克隆和表征，该菌株使用甲羟戊酸和非甲羟戊酸途径生成异戊烯二磷酸

T.Kuzuyama,M.Takagi,S.Takahashi and H.Seto: "Cloning and characterization of 1-deoxy-D-xylulose 5-phosphate synthase from Streptomyces sp.strain CL190 that uses both the mevalonate and nonmevalonate pathways for isopentenyl dephosphate biosynthesis"J.Bact

T.Kuzuyama、M.Takagi、S.Takahashi 和 H.Seto：“来自链霉菌菌株 CL190 的 1-脱氧-D-木酮糖 5-磷酸合酶的克隆和表征，该酶同时使用甲羟戊酸和非甲羟戊酸途径进行异戊烯基脱磷酸生物合成

S. Takahashi, T. Kuzuyama and H. Seto: "Purification, characterization, and cloning of a eubacterial 3-hydroxy-3-methylglutaryl coenzyme A reductase, a key enzyme involved in the biosynthesis of terpenoids"J. Bacteriol. 181. 1256-1263 (1999)

S. Takahashi、T. Kuzuyama 和 H. Seto：“真细菌 3-羟基-3-甲基戊二酰辅酶 A 还原酶的纯化、表征和克隆，该酶是萜类生物合成中涉及的关键酶”。